Ulcerative colitis (UC) and Crohn's disease, the two major forms of idiopathic Inflammatory Bowel Disease (IBD) in humans, are widespread and poorly understood disorders (Kirsner, J. B., et al., eds., Inflammatory Bowel Disease: 3rd ed., Lea and Febiger, Philadelphia (1988); Goldner, F. H., et al., Idiopathic Inflammatory Bowel Disease, in Stein, J. H., ed., Internal Medicine, Little Brown & Co., Boston, pp. 369-380 (1990); Cello, J. P., et al.. Ulcerative Colitis, in Sleisenger, M. H., et al.. eds., Gastrointestinal Disease: Pathophysiology Diagnosis Management, W. B. Saunders Co., Philadelphia, p. 1435 (1989)). The separation of these idiopathic diseases of unknown etiology from other forms of colitis and ileitis caused by infectious agents, drugs, or the solitary rectal ulcer syndrome and collagenous colitis is not always respected in the literature (Riddell, R. H., ed., Pathology of Drug-induced and Toxic Diseases, Churchill Livingstone, New York (1982)). The diagnosis of IBD of known and unknown etiology is often not only difficult but almost impossible, e.g., during serious local complications such as hemorrhage, toxic dilation, perforation, inflammatory polyps and strictures (Riddell, R. H., ed., Pathology of Drug-induced and Toxic Diseases, Churchill Livingstone, New York (1982)).
The pathology of ulcerative colitis usually refers to a more superficial mucosal disease in contrast to Crohn's disease with its deep, often transmucosal involvement and fissures (Riddell, R. H., ed., Pathology of Drug-induced and Toxic Diseases, Churchill Livingstone, New York (1982); Morrison, B. C., et al.. eds., Gastrointestinal Pathology, 2d ed., London (1979); Fenoglio-Preiser, C. M., et al., eds., Gastrointestinal Pathology: An Atlas and Text, Raven Press, New York (1989); Goldman, H., et al., Hum. Pathol. 13:981-1012 (1982)). Ulcerative colitis typically involves the rectum and extends proximally without intervening uninvolved "skip" areas which are usually the hallmarks of Crohn's disease. The histologic features of active ulcerative colitis include, beside the superficial ulcers, infiltration by inflammatory cells (e.g., mainly lymphocytes, plasma cells, variable number of neutrophils, eosinophils and mast cells) involving extensively the lamina propria. Crypt abscesses, i.e., aggregates of neutrophils near and invading the crypt epithelium are reliable indicators of activity, while depletion of mucin in goblet cells is a less frequent finding.
Despite all the controversy, most of the pathologists accept as "rule of thumb that well-formed, sarcoid-like granulomas are not part of the spectrum of UC and that if these are present, an alternative explanation must be found" (Riddell, R. H., ed., Pathology of Drug-induced and Toxic Diseases, Churchill Livingstone, New York (1982)). Foreign-body giant cells and collection of a few histiocytes, however, are often present because of rupture of crypt abscesses and spilling of mucin into the submucosa and eliciting of cellular reaction. Noncaseating granulomas, on the other hand, are often present in gut segments from Crohn's disease which is often also called granulomatous colitis.
Active ulcerative colitis is usually followed by resolving or quiescent forms of the disease. Alternatively, in the fulminant form of ulcerative colitis the deep ulcers extend into the muscularis propria, and the acute and chronic inflammatory cells also involve the submucosa and the true muscular layer in the vicinity of ulcers.
The etiology and pathogenesis of idiopathic IBD, as the name implies, are poorly understood. Numerous theories, however, implicate genetic predisposition, environmental factors, infectious agents and immunologic alterations (Kirsner, J. B., et al.. eds., Inflammatory Bowel Disease, 3rd ed., Lea and Febiger, Philadelphia (1988); Zipser, R. D., ed., Dig. Dis. Sci., 33 Suppl.:1S-87S (1988)). Previous understanding of the pathogenesis was limited to a three-stage process: (a) an irritant, which could be an immune process or infectious agent, activates (b) leukocytes which release enzymes such as proteases and inflammatory mediators such as histamine, serotonin and prostaglandins, and (c) these products cause edema, pain, heat and loss of function. However, as the overview of a recently held symposium on mediators of IBD concluded, this process is probably correct but it is far more complicated (Zipser, R. D., ed., Dig. Dis. Sci., 33 Suppl.:1S-87S (1988)).
The initial cell and tissue injury does not have to be a massive insult as subtle biochemical reactions such as generation of free radicals are also sufficient to initiate irreversible cell damage, especially if combined with periods of ischemia and reflow. The final and most potent mediator of this toxicity appears to be the hydroxyl radical derived from the iron-catalyzed interaction between superoxide and hydrogen peroxide (Grisham, M. B., et al., Dig. Dis. Sci. 33 Suppl.:6S-15S (1988)).
A working hypothesis was thus recently proposed stating that transient ischemic episodes in the intestine initiate a cascade of self-perpetuating cycles of reactive oxygen metabolites and free radicals. Tissue damage may be indirectly aggravated through inflammation as neutrophils generate substantial amounts of oxygen metabolites (Grisham, M. B., et al., Dig. Dis. Sci. 33 Suppl.:6S-15S (1988)). This possibility is in agreement with very recent data obtained on 27 patients with ulcerative colitis, 10 with acute bacterial diarrhea, and 20 healthy volunteers. These data indicate that the oxidative free radical generating capacity of polymorphonuclear leukocytes was markedly enhanced in patients with active ulcerative colitis as compared with controls and
patients in remission (Shiratora, Y., et al., Digestion 44:63-171 (1989)). These results indeed strongly suggest that increased free radical production by leukocytes could be related to the initial pathogenesis or aggravation of ulcerative colitis.
Results of studies performed during the last few years indicate that mediators of inflammation arise from cells other than leukocytes (Zipser, R. D., ed., Dig. Dis. Sci., 33 Suppl.:1S-87S (1988)). Fibroblasts, smooth muscle and other cells release prostaglandins which modulate blood flow and intestinal motility. The vascular endothelium and macrophages seem to be the source of interleukin-1 (IL-1) which is involved in the production of fever, lymphopenia and many metabolic changes (Dinarello, C. A., Dig. Dis. Sci. 33 Suppl.:25S-35S (1988)). A very recent report described an enhanced production of IL-IB by mononuclear cells isolated from mucosa of patients with active ulcerative colitis (Mahida, Y. R., et al.. Gut 30:835-838 (1989)). On the other hand, mononuclear cells from both forms of IBD generated less IL-2 than controls. Furthermore, increased number of IL-2 responsive killer cells or exacerbated reactivity to IL-2 was found, especially in Crohn's disease, indicating that reactivity to IL-2 distinguishes IBD from control intestinal mononuclear cells (Kusugami, K., et al., Gastroenterology 97:1-9 (1989)). .
Other mediators of inflammation in IBD include enhanced production of platelet-activating factor (PAF) during active disease and inhibition by sulfasalazine and prednisolone (Eliakim, R., et al., Gastroenterology 95:1167-1172 (1988)). Results from human and experimental IBD indicate an enhanced synthesis of eicosanoids such as prostaglandins, thromboxanes and leukotrienes (Schumert, R., et al., Dig. Dis. Sci. 33 Suppl.:58S-64S (1988)). These products may not only be involved in the pathogenesis of IBD but may have diagnostic value and serve as therapeutic targets. Specifically, the raised concentration of prostaglandin E2 in rectal dialysis fluid from patients with ulcerative colitis may identify patients with a risk of relapse (Lauritsen, K., et al., Gut 29:1316-1321 (1988)), while selective inhibition of leukotrienes may be a therapeutic strategy to reduce inflammation in IBD (Schumert, R., et al., Dig. Dis. Sci. 33 Suppl.:58S-64S (1988); Goetzl, E. J., et al., Dig. Dis. Sci. 33 Suppl.:36S-40S (1988); Allgayer, H., et al., Gastroenterology 96:1290-1300 (1989 )).
In addition to the established mediators, potential humoral mediators of inflammation may also be involved in the pathogenesis of IBD, e.g., tumor necrosis factor, growth factors, neuropeptides, lipoxins, and mast cell products (Zipser, R. D., ed., Dig. Dis. Sci., 33 Suppl.:IS-87S (1988); Shanahan, F., et al., Dig. Dis. Sci. 33 Suppl.:41S-49S (1988); Nast, C. C., et al., Dig. Dis. Sci 33 Suppl.:50S-57S (1988); Mayer, E. A., et al., Dig. Dis. Sci. 33 Suppl.:71S-77S (1988)). It is also possible that not only the number of inflammatory cells and their products are changed, but the number of receptors increase, such as the increased neutrophil receptors for and response to the proinflammatory peptide formyl-methionyl-leucyl-phenylalanine (FMLP) (Anton, P. A., et al., Gastroenterology 97:20-28 (1989)) and the adherence of leukocytes (Cason, J., et al., J. Clin. Pathol. 41:241-246 (1988)) in Crohn's disease.
The immunology of IBD remains a tantalizing and frustrating field (Hodgson, H. J. F., et al., Balliere's Clin. Gastroenterol. 1:531-542 (1987)). Despite very intensive investigations in patients and animal models of IBD, the basic question cannot be answered, i.e., whether ulcerative colitis and Crohn's disease are primarily manifestations of disordered immunity, or whether the immunologic abnormalities documented in idiopathic IBD secondary epiphenomena are generated during the process of disease (Hodgson, H. J. F., et al., Balliere's Clin. Gastroenterol. 1:531-542 (1987); Elson, C. O., The immunology of inflammatory bowel disease, in: Kirsner, J. B., et al., eds., Inflammatory Bowel Disease, Lea and Febiger, Philadelphia, pp. 97-164 (1988); MacDermott, R. P., et al., Adv. Exp. Med. Biol. 216A:335-344 (1987)). If the second possibility is correct, the nature of etiologic agent initiating the early cell and tissue injury remains unknown, and in addition, environmental and dietary factors as well as transmissible infectious agents should be considered (Calkins, B. M., et al., Epidemiol. Rev. 8:60-85 (1986); Myren, J., et al., Scand. J. Gastroenterol. 23 Suppl.:11-19 (1988); Mendelhoff, A. I., et al., The epidemiology of idiopathic inflammatory bowel disease, in Kirsner, J. B., et al., eds., Inflammatory Bowel Disease, Lea and Febiger, Philadelphia, pp. 3-34 (1988); Hodgson, H. J. F., et al., Balliere's Clin. Gastroenterol. 1:531-542 (1987); Elson, C. O., The immunology of inflammatory bowel disease, in: Kirsner, J. B., et al.. eds., Inflammatory Bowel Disease, Lea and Febiger, Philadelphia, pp. 97-164 (1988); MacDermott, R. P., et al., Adv. Exp. Med. Biol. 216A:335-344 (1987)). With these open dilemmas, immunologic studies of IBD have been concentrated on mechanisms that may be responsible for inflammation, irrespective of whether the pathogenesis of IBD has been initiated for immunologic reasons or in response to infection or other toxic agents (Hodgson, H. J. F., et al., Balliere's Clin. Gastroenterol. 1:531-542 (1987)).
The immunologic alterations in IBD are primarily autoimmune in nature, with colonic autoantibodies and lymphocyte-cytotoxicity directed against colonic epithelial cells. The immune response initially directed against bacteria in the gut could cross-react with intestinal epithelium due to antigenic similarity between these two targets. However, even the latest developments in the immunologic aspects of the pathogenesis of IBD cannot answer the basic question, i.e., whether the detected changes in humoral and cellular immunity reflect a primary defect or secondary response to injury.
Animal models are essential to study the etiology and pathogenesis of such a complex and probably multifactorial disease as idiopathic IBD. The criteria for an animal model of IBD and the insufficient quality and quantity of models have been reviewed repeatedly and extensively (Strober, W., Dig. Dis. Sci. 33 Suppl.:3S-1OS (1988); Beekan, W. L., Experimental inflammatory bowel disease, in: Kirsner, J. B., et al., eds., Inflammatory Bowel Disease, Lea and Febiger, Philadelphia, pp. 37-49 (1988)). The available animal models can be divided into naturally occurring and experimentally induced animal models. Unfortunately, only a few spontaneous and rarely occurring models of intestinal inflammation due to a genetic defect are available and most of these are not idiopathic but are induced by bacteria or other infectious agents (e.g., hyperplasia, crypt abscesses, ulcers in mice with Bacillus psyliformnis and hamster with "rod-shaped bacteria") (Strober, W., Dig. Dis. Sci. 33 Suppl.:3S-1OS (1988)). Rare forms of spontaneous ulcerative colitis and granulomatous enterocolitis also occur in rats and horses, respectively. Great expectations surrounded the initial reports on the potential use of marmosets and the cotton-top tamarin as a spontaneous model of ulcerative colitis and colonic adenocarcinomas (Clapp, N. K., et al., eds., Dig. Dis. Sci. 33 Suppl.:1S-158S (1988)). Unfortunately, multiple infections, the accompanying "wasting disease", the demanding conditions in husbandry and nutrition as well as the low survival rate (23-69%) of marmosets and tamarins contribute to the very limited availability of this grossly and microscopically good animal model of ulcerative colitis (Clapp, N. K., et al., eds., Dig. Dis. Sci. 33 Suppl.:1S-158S (1988)).
Experimentally induced animal models of ulcerative colitis are usually produced by exposure to toxic dietary substances, pharmacologic agents or other environmental chemicals, or by administration of materials derived from patients, or by manipulation of the animal's immune system (Strober, W., Dig. Dis. Sci. 33 Suppl.:3S-10S (1988); Beekan, W. L., Experimental inflammatory bowel disease, in: Kirsner, J. B., et al., eds., Inflammatory Bowel Disease, Lea and Febiger, Philadelphia, pp. 37-49 (1988); Onderdonk, A. B., Dig. Dis. Sci. 33 Suppl.:40S-44S (1988)). Although vascular changes, especially arterial occlusion and histamine administration, result in increased vascular permeability, mucosal necrosis and ulceration, the accompanying inflammation is very transient and cannot serve as a model of chronic ulcerative colitis (Beekan, W. L., Experimental inflammatory bowel disease, in: Kirsner, J. B., et al., eds., Inflammatory Bowel Disease, Lea and Febiger, Philadelphia, pp. 37-49 (1988); Krasna, I. H., et al., J. Pediat. Surg. 21:26-29 (1986)).
Neurogenic manipulation, especially the cholinergically induced hypermotility is also accompanied by limited colitis (Beekan, W. L., Experimental inflammatory bowel disease, in: Kirsner, J. B., et al., eds., Inflammatory Bowel Disease, Lea and Febiger, Philadelphia, pp. 37-49 (1988); Berger, R. L., Ann. Surg. 152:226 (1960)), and this might be an appropriate tool to study the stress-associated pathogenetic factors in ulcerative colitis (Szabo, S., Dig. Dis. Sci. 30 Suppl.:28S-31S (1985)). On the other hand, local administration of physical (e.g., hyperthermia) (Ger, R., et al., Dis. Colon Rectum 29:77-81 (986)) or chemical irritants (e.g., ethanol, acetic acid, formalin, detergents, hypertonic salt solutions and even nonsteroidal anti-inflammatory drugs) (Strober, W., Dig. Dis. Sci. 33 Suppl.:3S-10S (1988); Beekan, W. L., Experimental inflammatory bowel disease, in: Kirsner, J. B., et al., eds., Inflammatory Bowel Disease, Lea and Febiger, Philadelphia, pp. 37-49 (1988)) produce massive local tissue destruction which is often accompanied by very little or no inflammation. Furthermore, these nonspecific lesions usually heal relatively quickly and are not thus appropriate models of IBD.
Despite their limitations, the two most widely used models are the experimental colonic lesions produced by 2,4,6-trinitro-benzensulfonic acid (TNB) and carrageenan. Both models involve tissue destruction in the colon. Intrarectal administration of 5-30 mg of TNB in 0.25 ml of 50% ethanol in the rat produced dose-dependent colonic ulcers and inflammation which were maximal by gross and light microscopic examination at week, and by biochemical measurement of myeloperoxidase activity in the colon at 3-4 weeks (Morris, G. P., et al., Gastroenterology 96:795-803 (1989)). Histologically, the inflammatory infiltrate of mucosa and submucosa included polymorphonuclear leukocytes, lymphocytes, macrophages and connective tissue mast cells. Initially, massive edema and in the healing state (6-8 weeks) fibroblasts were also detected. Granulomas were seen in 57% of rats killed at 3 weeks.
In these experiments TNB acted as a hapten and ethanol apparently permitted its subepithelial penetration. Unfortunately, the TNB model is not reproducible without the nonspecific action of at least 30-40% ethanol (Morris, G. P., personal communication). Contrary to the results with rats, ethanol intake exerts a certain protection against IBD in humans. Furthermore, this rat model has been sensitive only to prevention (i.e., pretreatment) with 5-lipoxygenase inhibitors or leukotriene antagonists which, however, do not markedly influence the well developed state of disease (Morris, G. P., personal communication).
Carrageenan is a sulfated polygalactose (molecular weight above 100,000) widely used in the food industry and is considered safe for human use. Degraded forms of this polysaccharide (molecular weight 20,000-40,000) administered through drinking water induce ulcerative colitis in two weeks or later in experimental animals (Beekan, W. L., Experimental inflammatory bowel disease, in: Kirsner, J. B., et al., eds., Inflammatory Bowel Disease, Lea and Febiger, Philadelphia, pp. 37-49 (1988); Onderdonk, A. B., Dig. Dis. Sci. 33 Suppl.:40S-44S (1988); Benitz, K. F., et al., Food Cosmet. Toxicol. 11:565 (1973); Engster, M., et al., Toxicol. Appl. Pharmacol. 38:265 (1976)). In addition to ulcers, acute and chronic inflammation, macrophages laden with degraded carrageenan and suppressed phagocytosis are seen.
Carrageenan is toxic to monocytes and lymphocytes in vivo and after several days of culturing in vitro, and in vivo it enhances the lectin-dependent cellular cytotoxicity toward human cultured carcinoma cells (Beekan, W. L., Experimental inflammatory bowel disease, in: Kirsner, J. B., et al., eds., Inflammatory Bowel Disease, Lea and Febiger, Philadelphia, pp. 37-49 (1988)). A major advancement in the understanding of this experimental model of ulcerative colitis has been achieved after the recognition that B. vulgatus is required for the induction of carrageenan-induced colitis (Onderdonk, A. B., Dig. Dis. Sci. 33 Suppl.:40S-44S (1988); Onderdonk, A. B., et al., Am. J. Clin. Nutr. 32:285 (1979)). Furthermore, glandular atypism was noticed in rabbits on a 28-month study with carrageenan, and rats ingesting 1-10% carrageenan through diet develop colorectal adenomas and adenocarcinomas, providing thus an experimental link and tool to study the relationship of ulcerative colitis and carcinogenesis (Beekan, W. L., Experimental inflammatory bowel disease, in: Kirsner, J. B., et al., eds., Inflammatory Bowel Disease, Lea and Febiger, Philadelphia, pp. 37-49 (1988)). The carrageenan-induced colitis, however, exerts great variability in animal susceptibility and reproducibility, and requires several weeks or months for full development. In addition to carrageenan, the FMLP-induced experimental colonic lesions also represent a transition between chemically and cellularly induced animal models. This bacterial peptide activates and attracts neutrophils, and causes ulcers and inflammation in the rat ileum (VonRitter, C., et al., Gastroenterology 95:651-656 (1988); VonRitter, C., et al., Gastroenterology 96:811-816 (1989)). This new animal model, like the TNB, has not yet been extensively used.